The objective of the proposed research is to obtain information regarding the mechanism of ribosomal gene magnification in Drosophila. This will be done by examining the rRNA genes present on a bobbed chromosome after the first, second and seventh steps of magnification in order to determine what types of rDNA sequence changes have taken place. DNA from individual flies will be examined by transferring gel-fractionated, restriction enzyme-digested total DNA to nitrocellulose filters and hybridizing the filters with 32p-rDNA (Southern blots). After autoradiography, the hybridization patterns will be compared with rDNA patterns for the starting Xbb chromosome and for the Ybb chromosome used to magnify the Xbb ribosomal genes. By comparing the rDNA patterns it will be possible to determine if repeats in the magnified locus originate from the Xbb or Ybb nucleolus organizer or both. We will also determine whether a specific set of gene repeats is magnified in all flies or whether each individual magnifies a different set of repeats. Bobbed flies carrying a bb1 chromosome which have reverted to a bb+ (or nearly bb+) phenotype after one step of magnification will be examined and changes which have occurred during this rapid bobbed reversion will be compared with those which occur during step-wise magnification. We will also determine whether the BSY-bb chromosome increases its rDNA content during magnification, and whether reduction is always associated with magnification or whether it can occur independently of rDNA magnification. Results of these experiments will be used to distinguish among models for ribosomal gene increase during magnification. This information will contribute to our understanding of how specific gene amplifications in mammalian cells may occur, and will help us to understand genetic events which result in changes in gene copy number and in chromosome structure.